A Practical Guide to the Preparation of Liquid Crystal-Templated

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Review

A Practical Guide to the Preparation of Liquid Crystal-Templated Microparticles Xiaoguang Wang, Emre Bukusoglu, and Nicholas L. Abbott Chem. Mater., Just Accepted Manuscript • DOI: 10.1021/acs.chemmater.6b02668 • Publication Date (Web): 25 Aug 2016 Downloaded from http://pubs.acs.org on August 30, 2016

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Chemistry of Materials

A Practical Guide to the Preparation of Liquid CrystalTemplated Microparticles Xiaoguang Wang,†,* Emre Bukusoglu,†,* and Nicholas L. Abbott† †

Department of Chemical and Biological Engineering, University of Wisconsin-Madison, Madison, WI 53706.

ABSTRACT: We provide a practical guide to methods and protocols that use polymer networks templated from droplets of liquid crystal (LC) to synthesize micrometer-sized polymeric particles that are chemically patchy, anisometric in shape, possess anisotropic optical properties and/or are mesoporous. We describe a range of methods that permit the preparation of LC droplets (containing reactive monomers) as templates for polymerization, including formation of LC-in-water emulsions by either mechanical methods (e.g., vortexing), encapsulation in polymeric shells, or microfluidics. The relative merits of the methods, including ease of use and potential pitfalls, and the resulting droplet size distributions, are described. We also report a menu of approaches that can be used to control the internal configurations of the LC droplets, including changes in composition of the continuous solvent phases (e.g., addition of glycerol) and adsorption of surfactants or colloids at the interfaces of the LC droplets. Photo-polymerization of the LC droplets in bipolar, radial, axial or preradial configurations, and subsequent extraction of the nonreactive mesogens generates polymeric particles that have spindle, spherical, spherocylindrical or tear shapes, respectively. Finally, we describe how to characterize these polymeric particles, including their shape, internal structure, optical properties and porosity. The methods described in this paper, which provide access to complex microparticles with properties relevant to separation processes, drug delivery and optical devices, are general and versatile and can be readily developed further (e.g., by changing the choice of LC) to create an even greater diversity of microparticles.

INTRODUCTION Polymeric microparticles with complex shapes and internal structures possess properties and exhibit behaviors that are forming the basis of a range of promising technologies (e.g., for chemical separation processes,1-3 optical displays4,5 or drug delivery6,7). Motivated by this opportunity, we recently developed general and facile methods for the synthesis of anisometric polymeric microparticles by using droplets of thermotropic liquid crystal (LC) dispersed in an aqueous/glycerol continuous phase.8,9 The ordered LC within the droplets functions as a template for polymerization reactions, thus leading to the generation of micrometer-sized anisotropic polymeric networks. By preparing droplets containing mixtures of reactive and non-reactive mesogens with distinct internal organizations (so-called configurations, detailed below), photo-polymerizing the reactive mesogens, and then extracting the non-reactive mesogens, a range of polymeric particles with distinct shapes (spindle, sphere, spherocylinder and tear shape) can be generated as a function of the initial configuration of LC droplet (Figure 1). Inspection of the polarized light micrographs in Figure 1 also reveals that the microparticles synthesized from LC emulsions possess anisotropic optical properties (birefringence), indicating that they possess complex internal structures. The solid lines in the schematic illustrations shown in Figure 1 indicate the local orientation of the LC (the so-called director). In addition to preparing microparticles with distinct shapes, recent studies of these microparticles have revealed that control of the configuration of the LC within the templating

droplet, as well as the weight fraction of reactive mesogen, can be used to introduce nanoporosity.8 For droplets in so-called radial configurations, the mass density of spherical particles can be tuned from 0.2 - 0.6 g/cm3, as shown in Figure 2A. Electron micrographs of these templated microparticles revealed them to be mesoporous (Figure 2B and C). Nitrogen sorption isotherms confirmed the pore size and surface area in one set of microparticles to be 39 ± 16 nm and 476 m2/g, respectively.

Figure 1. Schematic illustrations and corresponding bright field (BF) and polarized light (PL) micrographs of (A) spindle-shaped, (B) spherical, (C) spherocylindrical or (D) tear-shaped polymeric microparticles templated from LC droplets. The orientations of the crossed polarizers in the PL micrographs are indicated by the white double-headed arrows. Scale bars: 5 µm.

Finally, we comment that the above-described methodology for microparticle synthesis also allows “patchy particles” to be

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prepared. In this extension of the synthetic methodology, solid colloids with desired surface chemistry are precisely positioned at topological defects present on the surfaces of the LC droplets (a detailed description of topological defects is given below).9-14 The surfaces of the solid colloids protrude from the LC droplets to define chemical patches. Figure 3A-D show spindle-shaped and tear-shaped microparticles, each with a single fluorescent polystyrene (PS) colloid positioned at one pole of the particle. Alternatively, arrays of colloids can be assembled on one half of the surface of a LC droplet (Janus-like particles), and the LC droplet polymerized to preserve the organization of the colloids (Figure 3E-H). This versatile method enables tailoring of the physical and chemical properties of the patches through the choice of material that constitutes the colloids.

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structure of the microparticles. The final section describes the synthesis and characterization of porous particles. We conclude by describing common problems encountered when using LC droplets to synthesize microparticles, and offer advice (learned the hard way!) on how to trouble-shoot them.

Figure 3. (A, C) Schematic illustrations and (B, D) corresponding BF and fluorescence micrographs of (A, B) spindle-shaped, or (C, D) tear-shaped microparticles with a single 1 µm-in-diameter fluorescent PS colloid adsorbed at the pole (indicated by white arrows). Janus-like microparticles viewed from (E, F) side, (G) top, and (H) bottom. The white dashed circles indicate the surface of the microparticle. Scale bars: 5 µm. Reproduced with permission.10

FORMATION OF LC DROPLETS

Figure 2. (A) Mass density of radial particles as a function of weight fraction of reactive monomer. Reproduced with permission.8 (B) Scanning electron microscope (SEM) and (C) transmission electron microscope (TEM) image of radial microparticles synthesized from 20 wt% reactive monomer followed by extraction of non-reactive mesogens.

Past papers detail the theory, physical principles and energetics that underlie the formation of the above-described complex particles.8-14 This Review, in contrast, aims to provide a practical guide to the enabling methods, procedures, and characterization techniques. In particular, we describe the relative merits of various experimental approaches and outline pitfalls and possible remedies. The Review is intended to be used by students and postdoctoral researchers without substantial prior experience experimenting with LCs. The remainder of this Review is organized into seven sections. The first section discusses methods for the formation of LC-inwater emulsions. In the second section, we describe common techniques that can be used to position colloids on the surfaces of LC droplets. Third, we highlight how to manipulate the configurations of LC droplets. The fourth and fifth sections detail procedures for photo-polymerization of LC droplets and subsequent extraction of non-reactive LC mesogens, respectively. In the sixth section, we introduce optical methods that can be used to characterize the shape and internal

In this section, we describe several methods that can be used to form LC droplets in ways that permit their subsequent use as templates for polymerization. As shown in Figure 4, the possible approaches include mechanical methods (vortexing, sonication, homogenization, gentle shearing by hand), encapsulation within polymeric multilayer capsules, and the use of microfluidics. Here we note that although all these procedures result in formation of LC droplets, each differ in terms of the droplet size distributions and ease of use, as summarized in Table 1. The non-reactive mesogen used in the examples described below is 4-cyano-4’-pentylbiphenyl (5CB), as shown in Scheme 1A. This mesogen forms a nematic LC phase at room temperature. The nematic-toisotropic phase transition temperature of 5CB is 35oC.

Scheme 1. Molecular structure of (A) 5CB and (B) RM257. (C) Schematic illustration showing the polymer network formed by RM257 in 5CB. The gray and white ellipsoids represent RM257 and 5CB, respectively. The black arrow indicates the local LC director.


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Chemistry of Materials to video S1 in the SI); (E) Encapsulating LC within polymeric microcapsules; (F) Preparation of LC droplets using microfluidics. Scale bars: 2 cm.

Figure 4. Methods for formation of LC droplets: (A) Vortexing LC in water (please insert direct link to video S1 in the Supporting Information); (B) Emulsifying LC using a homogenizer (please insert direct link to video S2 in the SI); (C) Sonication of LC in water (please insert direct link to video S1 in the SI); (D) Emulsifying LC in glycerol (please insert direct link

The preparation of LC droplets using mechanical methods (vortexing, sonication, homogenization, agitation by hand) starts with the addition of LC to an aqueous or glycerol phase (Figure 4A-D). Typically, 6 - 10 µL of LC added to 2 mL of a continuous aqueous/glycerol phase leads to an emulsion that is sufficiently dilute in LC droplets that minimal coalescence occurs prior to polymerization. Detailed descriptions of the mechanisms by which mechanical shearing leads to the breakup of the LC oil into micrometer-sized droplets can be found elsewhere.15 Here we focus on a discussion of the relative merits of the methods in the context of making LC droplets for polymerization. We first note that each of the methods shown in Figure 4A-D is quick to perform (a few minutes) but also leads to droplets that are polydisperse in size (see Table 1). The preferred method is influenced by the nature of the sample (e.g., presence of lipids, surfactants, polymers or colloids, viscosity of the continuous phase) and the desired range of LC droplet sizes. Although vortex mixers are ubiquitous in university laboratories and LC droplet sizes can be manipulated by varying the speed of vortexing (typically between 500 and 3,000 rpm), the method results in relatively large droplets sizes and it is not effective if the continuous phase is viscous (e.g., glycerol). Some of these limitations, however, can be overcome by the use of homogenizers and even with hand shearing. Homogenizers (Figure 4B) readily form droplets with diameters between 1 - 50 µm (e.g., using 5CB; 30 s at 6,500 rpm using a T25 digital ULTRA-TURRAX homogenizer equipped with a S25 N-10G dispersing element) whereas hand shearing forms droplets that are typically 10 µm or larger in size. For emulsification of very small volumes of LC, ultrasound generators are a good choice as they can be used with samples as small as ~100 µL. Typical sonication times were 1-5 mins, during which time sonication did not significantly change the temperature of the samples. Table 1 provides additional comments on these methods of emulsification. All the methods described above lead to LC droplets that are polydisperse in size. There exist, however, a couple of easily accessed methods that can be used to prepare LC droplets with well-defined sizes. First, it is possible to encapsulate LC droplets within polymeric capsules.16,17 This approach uses capsules prepared by layer-by-layer adsorption of polyelectrolytes onto silica (sacrificial) particles. Silica particles can be purchased with monodisperse sizes, thus enabling precise control of LC droplet sizes.16,17 In the example shown in Figure 4E, amine-functionalized silica particles are dispersed in water as sacrificial templates.18 Polymeric shells are prepared by sequential desorption of cationic and anionic polyelectrolytes (e.g., anionic poly(styrene sulfonate) (PSS) and cationic poly(allylamine hydrochloride) (PAH)) onto the sacrificial silica particles followed by etching of the silica using hydrogen fluoride (HF; HF is corrosive and extreme care should be taken while handling it). The resulting polymeric shells are transferred into an isotropic phase of mesogens and alcohol. Following evaporation of the alcohol (to form a LC phase within the capsule), the LC-filled capsules are extracted into an aqueous phase. Using this method, polymer-filled capsules with sizes ranging from 10.0 ± 0.22 µm to 0.7 ± 0.08 µm have been reported.18 Examples of hydrogen bonding capsules, which



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Method

Droplet size

Vortexer

~100 µm–1 mm

Sonicator

~100 nm – 10 µm

Homogenizer

~1 – 100 µm

Hand shearing in glycerol

~10 µm – 1 mm

Polymer capsules

~1 – 100 µm

Microfluidics

~10 – 100 µm

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Table 1. Methods for Formation of LC droplets. Comments Vortex speed and time can be varied to change the droplet size distribution Can decrease droplet size by adding surfactants in the aqueous phase Attention must be given to how to hold the vial during vortex (for consistency) Do not work well when using viscous liquids Allows the preparation of sample volumes as small as ~100 µL Droplets tend to be smaller than with other techniques Hard to achieve reproducible control of droplet size and distribution (the energy transferred to the sample within the bath appears to be strongly dependent on the location of the sample in the bath and details such as the amount of water in the bath) Can vary droplet sizes by varying the speed in the homogenizer More reproducible than vortexing Permits easy addition of colloids or amphiphiles Can be used with a viscous continuous phase The minimum volume of emulsion depends on the type of homogenizer tip (in milliliter range) Simple shear-induced fission of droplets. Quick and no complicated equipment (just a spatula!) Highly variable based on the energy level of the experimentalist Work well only when the continuous phases is viscous Droplet size depends on the size of silica particle templates Permit preparation of bulk quantities of droplets Relatively long and complex synthetic procedure Simple devices produce relatively small numbers of droplets (high throughput methods do exist if scale up is important) Excellent size control When using PDMS, lifetime of device is limited

can be readily disassembled after filling with LC (e.g., by a change in pH) have also been used to prepare “naked” LC droplets.19 Beyond providing access to samples of LC droplets with narrow size distributions, an advantage of this approach is that it is a bulk method and that can be used to prepare large quantities of droplets. Microfluidics provides an alternative to the above-described capsule-based methods for creating monodisperse LC droplets. The approach has been widely applied over the past decade for the preparation of monodisperse oil-in-water emulsions.20-28 Several designs have been reported, each differing in the nozzle geometry used to shear the oil phase into droplets.20-28 Microfluidic devices can be assembled from a variety of materials, including polydimethylsiloxane (PDMS), glass and silicon. An example of a microfluidic system prepared from PDMS is shown in Figure 4F. It was made by mixing 10:1 wt/wt PDMS base to initiator and curing the mixture for 3 hours at 70°C over a mold. The molded PDMS was subsequently cut into pieces, punched with tubing inlets, bonded to a microscope slide treated with an oxygen plasma, and then connected to inlet and outlet tubing. Within the device shown in Figure 4F, the oil phase passes through a narrow neck where it is sheared by an outer continuous phase into droplets. Microfluidic methods can readily generate droplets with diameters between 10 - 100 µm. Manipulation of the droplet size can be achieved by either modifying the channel geometry or by varying the flow rates of the LC and continuous phases. In practice, a limitation encountered with the use of small channels (< 10 µm) is that they tend to plug with solid particles/debris in the liquids. Also, when working with PDMS, it can be challenging to keep the channel walls hydrophilic and prevent adhesion of the emulsion droplets to the walls. This issue can be minimized by using devices within a few hours of their fabrication. We also note that the use of a

wide exit channel slows the flow and minimize the coalescence of droplets. We make three additional comments regarding the methods described above for the preparation of LC droplets. First, the use of glycerol as a continuous phase is a broadly useful strategy when using a homogenizer or sonicator. In general, the high viscosity of the glycerol leads to a substantial reduction in the LC droplet sizes (relative to that achieved when using water). LC droplets with sizes as small as a few hundred nanometers can be prepared. Second, LC droplet sizes can be reduced by the addition of surfactants. We have commonly used sodium dodecyl sulfate (SDS) in combination with vortexing to prepare emulsions with average sizes below 10 µm (please insert direct link to video S5 in the SI). Here we caution that the addition of the surfactant can also change the configuration of the LC within the droplet (details below), and that control of the concentration of the surfactant is important. As a rule of thumb, SDS concentrations in the micromolar range can facilitate emulsification yet are well below the concentrations at which the surfactant typically impacts the configurations of the LC droplets (~ 1 mM) or forms micelles (critical micelle concentration (CMC), which is around 8 mM for SDS). Third, addition of a depletant has been demonstrated to be an effective method for fractionating particles of a specific size from a polydisperse dispersion (since depletion attraction is a function of the particle size).29-31 A likely productive future direction would be to use depletion-induced fractionation techniques to obtain monodisperse droplets from polydisperse emulsions.

DECORATION OF LC DROPLETS WITH COLLOIDS As described in the Introduction, polymeric microparticles


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Configuration bipolar

radial

axial

preradial

Chemistry of Materials Table 2. Suggested methods to control configurations of LC droplets. Methods Comments Cannot be used for LC mixtures that do not exhibit Disperse LC in pure water planar anchoring at aqueous interfaces. Glycerol induces stronger planar anchoring as Disperse LC in glycerol compared to water.39 40 Adsorb polymers that cause planar anchoring (e.g., PVA ) on The anchoring energy of the interface can be the surface of LC droplets manipulated by varying the polymer concentration. Adsorb synthetic surfactants (e.g., SDS34,41,42) and lipids (e.g., Requires careful control of the concentration of amphiphile or salt. The molecular architecture of the DLPC,41,43 Lipid A44,45) on the surfaces of LC droplets amphiphile can play an important a role in 46 Addition of salts at high pH (e.g., 1 M NaCl at pH 13) determining the ordering of the LC. Adsorb intermediate concentrations of surfactants on the Precise control over the droplet size may be required surface of LC droplets (e.g., 0.2 mM SDS34) to achieve desired configurations. Dope LCs with molecules that induce tilt in surface anchoring Adsorb intermediate concentrations of surfactants on the Precise control over the droplet size may be required surface of LC droplets (e.g., 0.6 mM SDS34) to achieve desired configurations. Dope LCs with molecules that induce tilt of surface anchoring Colloids adsorbed on droplets pin point defect at the Adsorption of colloids in the presence of surfactants8,11,12 surface of the droplets, resulting in LC droplets exhibiting homogeneous configurations.

with chemical patches can be prepared by the adsorption of solid colloids onto the surfaces of LC droplets followed by photo-polymerization.8-10 We have typically adsorbed colloids onto the surfaces of LC droplets by adding an aqueous colloidal dispersion to a preformed LC-in-water emulsion and then homogenizing the system at 6,500 rpm using a ULTRATURRAX homogenizer. The average number of colloids adsorbed onto the LC droplets increases with the duration of the homogenization. Once adsorbed onto the LC droplets, the colloids migrate to the locations of topological defects formed at the surfaces of the LC droplets (see below for details). The localization of the colloids at defects is independent of the surface properties of the colloids.9 The surface charge of the colloids, however, plays an important role in the adsorption of the colloids onto the LC droplets (the surface charge of the LC droplets is negative). We comment here that in the absence of homogenization, very few PS or silica colloids adsorb onto the surface LC droplets in pure water due to electrostatic repulsion. Adsorption of colloids on LC droplets can be enhanced by increasing the ionic strength of the continuous aqueous phase.

MANIPULATION OF THE INTERNAL CONFIGURATIONS OF LC DROPLETS When nematic LCs are confined within micrometer-sized droplets, configurations with distinct optical signatures are exhibited by the LC. The configurations reflect the energetic contributions of the elastic deformation of the LC, the orientation-dependent interfacial energy of the LC at the droplet surface (so-called surface anchoring), and the energetic cost of creating the cores of topological defects (nanoscopic regions of LC where the elastic energy density induces local disorder in the LC).32,33 When using LC droplets as templates for polymerization, the configuration of the LC droplets is most readily manipulated by tuning the surface anchoring of the LC using surfactants adsorption. Common configurations of nematic LC droplets are shown in the left column of Table 2. When LC is aligned tangential (locally parallel) to the droplet surface, a bipolar configuration is observed, in which two diametrically opposed surface point defects (so-called

boojums) are present at the poles.18,32-38 By tilting the surface anchoring of the LC at the droplet surface (away from tangential), an axial configuration is obtained, as evidenced by the disappearance of the two boojums of the bipolar droplet and the simultaneous appearance of a disclination ring near the droplet equator.32-34,36 With further tilting of LCs at the droplet surface (towards the radial orientation), this ring defect moves towards one pole and shrinks to a surface point defect, leading to a preradial configuration.32-34,36 Finally, when the LCs are perpendicular at the droplet surface, a point defect forms at the droplet center, leading to a radial configuration.32-34,36 Table 2 lists a range of strategies that can be used to tune the interfacial interactions of LCs to obtain near-homogeneous populations of bipolar, axial, preradial and radial LC droplets. As mentioned in the previous section, LC droplets with chemical patches can be synthesized by the adsorption of colloids onto the surfaces of the droplets. Previous studies have shown that the configurations of LC droplets influence the positioning of the colloids.9-13,35 For example, colloids will locate at both surface defects of bipolar droplets (two diametrically opposite boojums) whereas they will localize to the single surface defects of preradial droplets. Past studies have also found that adsorbed colloids can alter the configurations of LC droplets.8,10,12 For example, at a concentration of 1 mM SDS, LC droplets free of surfaceadsorbed colloids adopt radial configurations, whereas droplets with adsorbed colloids exhibited preradial configurations. This result shows that colloids can pin topological defects to droplet surfaces. We comment here that reversible switching of the internal configurations of LC droplets can be used to sweep colloids from multiple local surface energetic minima (i.e., two boojums per bipolar LC droplet) to a single face of the LC droplet surface, leading to homogeneous populations of dipolar patchy microdroplets.10 The positions of the adsorbed colloids can be preserved by photo-polymerization, as will be described in the next section. We end this section by commenting on the range of LC droplet sizes that can be used with the methods listed in Table 2. The elastic energy of a LC droplet scales with the radius of



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Problem Unwanted/unexpected shapes of microparticles

Table 3. Trouble-shooting. Possible causes Non-uniform surface anchoring of LCs due to lack of mixing of glycerol-water mixture, creaming at air-liquid interface, etc.

LC droplets adhere to walls of microfluidic channel

Hydrophobic walls of the microfluidic device.

LC droplets coalesce at the outlet of the microfluidic device Polymer aggregates observed within the LC droplets after polymerization

Motion of droplets at the channel outlet causes coalescence. Reactive mesogen mixture is contaminated or prepolymerized, LC-reactive mesogen mixture is not well mixed.

Fluorescent signal in LC droplets with surface-adsorbed colloids

Fluorophores dissolve into LC.

the droplets (KR, where K is the elastic constant of LCs and R is the radius of the LC droplets) and the surface anchoring energy scales with the square of R (WR2, in which W is the surface anchoring strength). These two free energies lead to the expectation that droplets with R > K/W will exhibit a variety of configurations that satisfy surface anchoring, and thus can be used with the methods in Table 2.18,32,33,47 A typical value for K/W for a low molecular weight LC is around 1 µm.

PHOTO-POLYMERIZATION DROPLETS

OF

LC

In our approach to the synthesis of complex polymeric particles, the various LC droplet configurations reported above serve as templates for photo-polymerization. This leads to polymeric networks that are structured by the nematic order of the LC droplets, as shown in Scheme 1C. To this end, we have generally used mixtures of 5CB and the reactive mesogen 4(3-acryloyloxypropyloxy) benzoic acid 2-methyl-1,4phenylene ester (RM257; Scheme 1B). RM257 is a bifunctional cross-linker. RM257 forms, upon photopolymerization, a highly cross-linked polymer network. Photopolymerization of aqueous emulsions of the LC droplets in their desired configuration can be performed by using a UV lamp (365 nm) that delivers 2.5 mW/cm2 at a distance of 5 cm from the light source. We have typically exposed the emulsions to UV light in glass vials for 30 min, gently mixing the emulsions every 10 min during the polymerization. This procedure leads to spherical microparticles comprised of a crosslinked polymer network swollen with non-reactive mesogens. Importantly, polymerization of RM257 within 5CB occurs with minimal perturbation to the anisotropic internal ordering of the confined LC.8 The above-described procedures can also be used to prepare polymerized Janus-like LC droplets with colloids adsorbed on one hemisphere of the droplets, as shown in Figure 3E-H. Photopolymerization of LC droplets is simple to perform, but we have encountered conditions/procedures that lead to failed syntheses. Experimental challenges and remedies are summarized in Table 3. For example, we found it necessary to use glycerol instead of water as the continuous phase when preparing bipolar droplets of RM257/5CB because the RM257 causes the LC to assume a tilted orientation at the droplet surface in pure water (glycerol induces strong tangential anchoring; see Table 2). If the configuration of the LC droplet observed after photo-polymerization differs from the initial configuration, a likely cause of failure is creaming of particles at the air—

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Solution Mix the aqueous glycerol mixture well, mix frequently during polymerization. Treat the channel walls to make them hydrophilic, such as via oxygen plasma etching. Make the exit channel wider to slow down the LC droplets. Make fresh samples and mix well. Prevent reactive mesogens from exposure to UV-light and/or heat source. Perform the experiments using freshly prepared reagents. Use fluorophores that are not soluble in LC.

glycerol interface (glycerol is more dense than the LC mixture), which leads to non-uniform surface anchoring (the LC adopts a homeotropic orientation at air—LC interfaces). To remedy this problem, the emulsion should be mixed frequently during photo-polymerization. We also comment that we sometimes observed aggregates within the LC droplets after polymerization (or even before exposure to UV). This was caused by either RM257 polymerized by ambient light exposure, or poor mixing of the LC/reactive mesogen mixture. In general, these problems can be avoided by using freshly prepared and well-mixed reagents, and avoiding exposure of the reagents to heat or UV light prior to use.

EXTRACTION OF LCS The photo-polymerization procedure reported in the previous section leads to the formation of spherical LC droplets containing a polymeric network. The final step in the preparation of the polymeric microparticles is extraction of the non-reactive mesogen (5CB) from the polymerized droplets. This step results in the shrinkage of the anisotropic polymer network into a solid polymeric particle. The shape of the microparticle is dictated by the configuration of the LC droplet used to synthesize the polymer network, as shown in Scheme 2. To extract the unreacted mesogen following photopolymerization, we added 0.2 mL of polymerized emulsion to 1.8 mL of ethanol. The diluted emulsion was then placed into a 2 mL polypropylene microcentrifuge tube, and centrifuged for 20 min at 4,500 rpm. After centrifugation, 1.8 mL of the supernatant was removed, and 1.8 mL of ethanol was added to each tube to re-disperse the pellet. This extraction step was repeated three times. Following extraction, 500 µL of water was added to re-disperse the pellet of polymeric microparticles. We note here that sonication should not be used to re-disperse the pellet as the polymeric microparticles can be fragmented by this process. Upon extraction of the non-reactive mesogens, the polymerized LC droplets shrink preferentially in a direction perpendicular to the local director.8,9 For example, polymerized bipolar droplets shrink preferentially in a direction perpendicular to the line joining the poles of the bipolar droplets, thus yielding spindle-shaped microparticles, as shown in Figure 1A. When the LC droplets have surfaceadsorbed colloids, we observed the positions of the colloids to be preserved during polymerization, thus yielding polymeric particles with both chemical patches and anisometric shapes. For example, spindle-shaped and tear-shaped microparticles


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with fluorescent PS colloids adsorbed at one pole of the particles can be obtained by photo-polymerizing bipolar droplets and preradial droplets followed by extraction of 5CB, respectively, as shown in Figure 3A-D and Scheme 2B.

symmetric (scheme in Figure 1B). To determine if the polymeric network is perpendicular or parallel to the direction of the propagating incident light, the polarizers (or the sample) are rotated 45°. Rotation of the dark texture together with the polarizers indicates a perpendicular alignment of the polymer network relative to the direction of the propagating light.

CHARACTERIZATION OF POROSITY OF TEMPLATED POLYMERIC PARTICLES

Scheme 2. Synthesis of (A) anisometric and (B) patchy polymeric microparticles templated from LC droplets.

OPTICAL CHARACTERIZATION OF THE SHAPE AND INTERNAL STRUCTURE OF POLYMERIC PARTICLES Characterization of the shape and internal structure of microparticles templated from LC droplets is most easily achieved using a combination of optical and electron microscopy. To characterize the shapes of the microparticles, we have typically used bright field (BF) microscopy in transmission mode (no polarizers in the optical path), as illustrated by the images shown in Figure 1. For particles in sub-micrometer sizes, SEM is a good candidate for characterization (see Figure 2B). When colloids are adsorbed to the LC droplets (to create chemical patches), we have used either BF microscopy or fluorescence microscopy (if the colloids are labelled with a fluorophore). The images reported in this paper (Figure 3) were obtained using a transmission mode of Olympus IX71 microscope, in which the samples were illuminated from bottom and observed from a camera above the sample. Polarized light (PL) microscopy is a very useful technique for charactering the internal structure of the LC droplets and polymeric microparticles templated from the LC droplets. The LCs and the polymerized RM257 network possess orientational order which gives rise to anisotropic optical properties (i.e., one refractive index perpendicular to the LC director (ordinary refractive index no) and the other one parallel to the director (extraordinary refractive index, ne)). The theory and practice of PL microscopy has been detailed in prior publications.32,48 Here we make a few comments relevant to the interpretation of PL micrographs of polymeric microparticles presented in this paper. First, we note that, when viewed through crossed-polars, the polymeric network within a microparticle will exhibit a dark optical appearance when the polyRM257 is oriented either parallel to the direction of the propagation of incident light through the particle or parallel/perpendicular to the crossed-polars.48 Here we use particles exhibiting a radial configuration to illustrate how to use the optical appearance of the microparticles to obtain insights into the internal structure. As shown in Figure 1B (bottom image), the microparticle exhibits a centered, dark cross aligned with the orientations of the polarizers, implying that the orientation of the polymer network is spherically

We observed the extraction of 5CB from polymerized radial droplets to result in only minimal shrinkage of the particles (and preservation of the spherical shape). Because very little shrinkage took place, the mass density of the spherical particles could be tuned from 0.2-0.6 g/cm3 with increase in mass fraction of RM257 from 10-40%, as shown in Figure 2A. To characterize the accompanying change in porosity of the spherical particles, two characterization methods, transmission electron microscope (TEM) and nitrogen sorption isotherm, can be used.8 To prepare samples for TEM imaging, spherical particles are mixed with 1 wt% PVA (in water) and then dried. Subsequently, the particle-embedded PVA films are cut into 80 nm-thick slices using an ultramicrotome and placed on TEM grids for imaging. As shown in Figure 2C, the pores of spherical particles templated from 20 wt% RM257 are in the range of 10-100 nm. To characterize the porosity, ~ 0.1 g of dry spherical particles are placed in a Micromeritics ASAP 2020 Accelerated Surface Area and Porosimetry system. The particles are degassed under vacuum at 423 K. Next, nitrogen is used at 77 K to obtain the pressure-adsorbed mass density isotherms of spherical polymeric particles. Finally, the average pore size of spherical particles can be estimated from the isotherm by using Barrett-Joyner-Halenda (BJH) theory (see SI for details).49 For spherical particles synthesized from 20 wt% RM257 in 5CB, the volume-average pore diameter was measured to be 39 ± 16 nm, consistent with the results by using TEM.

CONCLUSIONS The methods and protocols described in this Review enable the synthesis of a wide range of patchy, anisometric microparticles with complex internal structure and porosity. This diversity reflects the various configurations that LC droplets can assume as templates for photo-polymerization. Accordingly, the Review is focused on describing methods to prepare LC emulsions with different sizes and internal configurations. Upon photo-polymerization of the reactive monomer and extraction of the non-reactive mesogens, LC droplets exhibiting different configurations template the formation of micrometer-sized anisometric and patchy polymeric particles. Overall, the methods reported in this paper have the advantage that they do not involve the use of complex instrumentation or procedures. The procedures can be optimized to enable the synthesis of microparticles for a range of potential applications, including intracellular delivery, photonics, separations and biocatalysis. They can also be adapted to other types of LC phases, such as cholesteric phases or blue phases,50-54 and thus expand further the range of microparticles that can be synthesized using LC droplets. We end by noting four directions that we judge to be potentially productive for future inquiry: (i) It is likely that the internal pore structures of the porous particles described in this Review are anisotropic. Characterization of the pore structures



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and associated mass transport properties of the particles is needed. (ii) We believe it should be possible to tailor the chemical functionality of the pores of porous particles by, for example, copolymerization of RM257 and monomers having acidic or basic functional groups. (iii) The anisometric particles synthesized by templating from LC droplets might be used to explore clustering and gelation phenomena in colloidal dispersions comprised of non-spherical particles. (iv) As described earlier in the Review, additional experiments are needed to combine our methods with scalable techniques that can be used to prepare monodisperse emulsions (e.g., depletion-induced fractionation method29-31).

ASSOCIATED CONTENT Supporting Information. The Supporting Information is available free of charge via the Internet at http://pubs.acs.org. SI includes: Characterization of porosity of microparticles using nitrogen sorption isotherms; Videos S1-S5 of formation of LC emulsions using vortexer, sonicator, homogenizer or hand mixing.

AUTHOR INFORMATION Corresponding Author *[email protected] and [email protected]

Notes The authors declare no competing financial interest.

ACKNOWLEDGMENT This work was supported by the National Science Foundation (under awards DMR-1121288 (MRSEC) and CBET-1263970), the Army Research Office (W911-NF-11-1-0251 and W911-NF14-1-0140), and the National Institutes of Health (AI092004).

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Table of Contents (TOC)


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A Practical Guide to the Preparation of Liquid Crystal-Templated

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